Abstracts

RATIONALE: Reduction in the effect of GABAergic agents as status epilepticus (SE) proceeds is well known (Treiman et al, 1998; Kapur & Macdonald, 1997). We hypothesisze that a reduction in post-synaptic GABA[sub]A[/sub] receptors occurs during SE with physiologic consequences on GABA inhibition. The participants should appreciate a relation between status epilepticus, GABA[sub]A[/sub] receptor internalization and altered GABA[sub]A[/sub] synaptic physiology at the end of the presentation.
METHODS: Immunocytochemistry was performed on 6-8wk old Wistar rats who had intra-cardiac perfusion with paraformaldehyde after 1 hour of lithium (3 mEq/kg ip) [ndash] pilocarpine (40 mg/kg ip) induced SE. Brain slices thru hippocampus were double labeled with antibodies for [beta]2/[beta]3 subunits and synaptophysin and viewed by confocal micropscopy. Visualized granule whole cell patch-clamp recordings of mIPSCs (using tetrodotoxin) were performed in hippocampal slices from SE and control animals.
RESULTS: GABA[sub]A[/sub] receptor internalization occurs by 1 hour of SE. Dentate granule cells, hilar interneurons and CA3a pyramids demonstrate colocalization of the [beta]2/[beta]3 subunits with synaptophysin on the surface of soma and proximal dendrites in controls with internalization of those subunits in SE. Initial results show 12% of [beta]2/[beta]3 subunits are internalized in controls compared to 47% in rats in SE for 1 hr with the lithium-pilocarpine model. mIPSCs recorded from dentate granule cells in acute slices prepared 1 hour after pilocarpine SE show a decrease peak amplitude to 61.8([plusminus]11.9)% of control values. With an intermittent perforant path stimulation (PPS) model of SE (20 Hz 10 sec/min and 2 Hz continuous pulses, .1ms duration, 20V), we noted sustained loss of inhibition of paired-pulse responses after even brief periods of stimulation. With only 1-3 min of stimulation, loss of inhibition lasted 43 ([plusminus]15) min in the dentate [italic]in vivo[/italic] [as measured by increase paired-pulse ratio (P2/P1 ISI 40ms) from .25 ([plusminus] .27) pre- to 1.02 ([plusminus] .18) post-stim; (p[lt] .001)] and in hippocampal slice preparations (where it is mimicked by low dose GABA[sub]A[/sub] antagonists).
CONCLUSIONS: The immunocytochemical findings of GABA[sub]A[/sub] receptor internalization and peak amplitude decrease of mIPSCs after 1 hour of SE suggest that a reduction of cell surface GABA[sub]A[/sub] receptor numbers has early effects on synaptic inhibition and may herald the transition to self-sustaining SE. In addition, the sustained loss of paired-pulse inhibition to very brief perforant path simulation suggests a profound and rapid onset of alteration in GABAergic inhibition to mild convulsant stimuli. The early effects on inhibition may serve as an initial events or triggers to seizure onset.
[Supported by: a VA Career Development Award (to D.N.) and by Research Grant N13515 from NINDS (to C.W., H.L.)]